Inhibition of tartar deposition by polyanionic/hydrophobic peptides and derivatives thereof which have a clustered block copolymer structure

ABSTRACT

A method of treating a tooth so as to inhibit deposition of a mineral thereon, which comprises contacting the tooth with a mineral deposition inhibitory effective amount of a poly-amino acid compound which has the structure: 
     
         poly(X).sub.n poly(Y).sub.m 
    
     where each X is a residue independently selected from the group consisting of aspartic acid, glutamic acid, phosphoserine, phosphohomoserine, phosphotyrosine, and phosphothreonine, 
     each Y is independently a residue selected from the group consisting of alanine, leucine, isoleucine, valine and glycine, 
     n is 2 to 60, 
     m is 2 to 60, and 
     n+m&lt;5, 
      and wherein poly (X) n  may contain up to 10% of the Y residues and poly (Y) m  may contain up to 10% of the X residues, and salts thereof, 
     in combination with an orally acceptable vehicle compatible with said compound.

Work for the present invention was supported in part by grants from theAlabama Research Institute, The National Science Foundation, and theNational Oceanic and Atmospheric Administration (Sea Grant).

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to new peptides and polypeptides that arepowerful inhibitors of tartar (dental calculus) formation.

2. Discussion of Background

Biological mineralization is a fundamental process in nature. Formationof bones and teeth from calcium phosphate and formation of shells andreefs from calcium carbonate are but two examples of this process.

Unfortunately, mineral deposits also frequently occur where they are notwanted. In the body, mineral deposition may contribute to dental plaque,hardening of the arteries, various organ stones, and failure ofprosthetic devices like implanted heart valves. In the marineenvironment, the biomineral structures may cause problems as in the caseof barnacles growing on the hulls of ships, adding extra bulk andcreating drag. In industry, mineral scale forms on surfaces of coolingtowers and other devices preventing their proper operation as heatexchangers and frequently promoting localized corrosion.

Because of the problems associated with unwanted mineral deposition,much effort has been devoted to finding mineralization inhibitors,particularly in industry, that might be used to prevent harmful mineralformation.

Molecules for prevention of mineral deposition have ranged from simpleions like Mg⁺² (Pytkowicz, R. M., J. Geol. 73, 196-199 (1965)) and PO₄³⁻ or pyrophosphate (Simkiss, K., Biol. Rev. 39, 487-505 (1964)) to morecomplex organic materials. Inhibition by simple ions is based on theability of these ions to interfere with the orderly formation of thecrystalline lattice of the mineral, such as CaCO₃. In addition,phosphate and polyphosphates have the property of protecting metallicsurfaces by forming very thin films that cover potential sites ofcorrosion on the surfaces.

Phosphonates were introduced in the late 1960's (Ralston, U.S. Pat. No.3,336,221 (1967)). These are small organic molecules with PO³ groupsattached directly to a central carbon atom via a covalent bond tophosphorus. The phosphonates are very effective inhibitors ofcrystallization that work by adsorbing to crystal surfaces.Hydroxyethylidene diphosphonate (HEDP) is perhaps the most widely usedphosphonate, still among the most powerful inhibitors of CaCO₃ formationknown.

Use of phosphonates has some disadvantages though. For example,phosphonates can be degraded during chlorination which in turn may leadto elevated phosphates and associated phosphate scales. Phosphonatesthemselves may also precipitate under common operating conditions. HEDPis an exceptionally effective inhibitor of crystal nucleation on aweight basis as shown by its effect on lengthening the induction periodprior to crystal growth, but it is not at all effective at inhibitingcrystal growth after it begins, (Sikes and Wheeler, CHEMTECH, in press(1987)).

More recently, as a result of continuing efforts to identify betterinhibitors, polyacrylate and other polyanionic materials have beenidentified (Rohm and Haas Company, Technical Bulletin CS-513A (1985),Fong and Kowalski, U.S. Pat. No. 4,546,156 (1985)). In the 1980's,antiscaling and anticorrosion technology has been based increasingly onuse of synthetic polymers under alkaline conditions. The current trendin synthetic polymers for water treatment is the use of randomcopolymers or terpolymers with alternating side groups of COO- withgroups like OH, CH₃, PO₃ ²⁻, SO₃ ²⁻ etc.

A new approach to identifying mineralization inhibitors was disclosed bySikes and Wheeler U.S. Pat. Nos. 4,534,881 (1985); 4,585,560 (1986); and4,603,006 (1986) and Wheeler and Sikes, U.S. Pat. No. 4,587,021 (1986).In these patents, it was disclosed that proteins and polysaccharidesextracted from biological minerals like oyster shells are stronginhibitors of crystallization. By studying the structure of the naturalmolecules, particularly the proteins, clues to the chemical naturerequired for activity of synthetic polyamino acids were obtained. Basedon this, certain polyanionic polyamino acids including random copolymersof negatively charged residues and nonionic residues were identified asuseful inhibitors.

Some other patents related to this invention are the following:

Gaffar, U.S. Pat. No. 4,339,431, discloses copolymers of glutamic acidand alanine which are used as anticalculus agents in dentifrices ormouthwashes. The copolymers disclosed therein are random copolymers.

Segrest et al, U.S. Pat. No. 4,643,988, discloses polypeptides having anon-clustered arrangement of anionic and hydrophic amino acids, whichwere used for treatment of atherosclerosis (perhaps by preventingdeposition of certain minerals).

Buck, U.S. Pat. No. 4,362,713, is directed to compositions and methodsfor preventing attachment of dental plaque to the surface of teeth bythe use of salts of certain maleic acid copolymers.

In spite of the above approaches to solving the problems of unwantedmineral deposition, there remains a strong need for a new and morepowerful inhibitors of mineral deposition which could be used in thebody, in a marine environment, or industrially.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: Solid phase peptide synthesis applicable to preparing thepresent peptides and polypeptides.

FIG. 2: Data obtained from a pH-drift crystallization assay.

FIG. 3: Data obtained from a calcium phosphate crystallization assay.

FIG. 4: Graphs showing the initial growth rate of CaCO₃ crystals (A),the growth rate of crystals after matrix addition (B), and cryptedgrowth rate (C) after the addition of an inhibitor as determined in apH-stat.

FIG. 5: Data obtained from a CaCO₃ crystallization, constant compositionassay.

FIG. 6: The effects of synthetic peptides on CaCO₃ crystallization(CaCO₃ pH-drift assay).

FIG. 7: Inhibition of CaCO₃ crystal growth by synthetic peptides;constant composition seeded crystal assay.

SUMMARY OF THE INVENTION

Accordingly, it is an object of the present invention to provide new andimproved compositions for inhibiting mineral deposition.

It is another object of the present invention to provide compositionsfor prevention of formation of tartar on teeth.

It is yet another object of the present invention to provide for methodsof prevention of the above-mentioned types of mineralization.

These and other objects of the present invention which will hereinafterbecome more readily apparent, have been accomplished by discovering thatpolypeptide materials that are polyanionic on one end of the moleculeand hydrophobic on the other end have the capability of stronglyinhibiting tartar deposition on teeth. These materials have thefollowing general formula:

    poly (X).sub.n poly (Y).sub.m

where each X independently is aspartic acid, glutamic acid,phosphoserine, phosphohomoserine, phosphotyrosine, or phosphothreonine,

each Y independently is alanine, leucine, isoleucine, valine, glycine orother nonpolar, amino acid residues,

n is 2 to 60,

m is 2 to 60, and

n +m is ≧5,

and wherein poly (X)_(n) may contain up to 10% of the Y residues andpoly (Y)_(m) may contain up to 10% of the X residues, and salts thereof.

The present invention is directed to compositions containing thesematerials, such as dentifrices and mouthwashes for oral application, aswell as methods of treating teeth by use of these compositions.

A more complete appreciation of the invention and many of the attendantadvantages thereof, will be readily perceived as the same becomes betterunderstood by reference to the following detailed description ofpreferred embodiments thereof.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

An interest in understanding better the chemical requirements foractivity as inhibitors of mineralization prompted the inventors toidentify new chemical structures of polypeptides that have surprisinglyenhanced inhibitory activity. As a basis for this work, the inventorsfurther elucidated the chemical structure of oyster shell proteins. Theyalso took into consideration the structure of certain salivary proteinsthat inhibit crystallization (Hay, D. I. et al, Inorg. Persp. Biol. Med.2, 271-285 (1979) and Calcif. Tiss. Int. 40, 125-132 (1987)).

The polypeptides of the present invention are synthetic polypeptideswhich possess clustered polyanionic amino acids at one end of themolecule and clustered non-ionic partly hydrophobic amino acids at theother end of the molecule. Preferably, the anionic amino acids arelocated at the C-terminus whereas the non-ionic amino acids are locatedat the N-terminus, although the opposite arrangement is alsocontemplated. Until the present invention, no one recognized thatsynthetic polypeptides having such a structure could be potentinhibitors of mineralization.

Although not wishing to be limited by a hypothesis as to the mechanismof action of the present polypeptides, the following is presented as apossible mechanism of action. The polyanionic region might stick tocrystal surfaces, blocking growth there, while the hydrophobic regionmight extend from the surface and disrupt diffusion of lattice ions toother growth sites. This hypothesis is referred as the PH orpolyanion/hydrophobe hypothesis. It is novel from the standpoint thatcrystal formation generally is thought to be limited not by diffusionbut rather by the rate of surface reactions in which adsorbed ionsbecome incorporated into crystal growth sites (Nancollas, G. H. and M.M. Reddy, J. Coll. Inter. Sci. 37, 824-830 (1971); Nancollas, G. H.,Adv. Coll. Inter. Sci. 10, 215-252 (1979)). This conclusion is based onthe calculated energy of activation of crystallization for CaCO₃formation of 11.0 kcal/mole which is thought to be too high for adiffusion-controlled mechanism (Howard, J. R. et al, Trans. Faraday Soc.56,278 (1960)), and on the observed lack of effect of the rate ofstirring on rates of seeded crystal growth. Although this may be thecase under some conditions, the inventors have often noticed thatstirring is quite important and in fact must be carefully controlled toachieve reproducibility in many crystallization assays. They suggestthat diffusion can be rate-limiting. In addition, studies of the effectsof polyacrylates on the inhibition of crystallization suggested that lowmolecular weight polymers have the greatest effect on a weight basis(Rohm and Haas, op. cit.).

Based on the above information, the present inventors hypothesized thatlow molecular weight polypeptides (e.g. 500-5000 daltons) with apolyanionic/hydrophobic arrangement could have useful properties asinhibitors of mineralization.

Higher molecular weight polypeptides (e.g. 5000-10,000 daltons) havingthe clustered polyanionic/clustered hydrophobic structure of theabove-described lower molecular weight polypeptides are also expected tohave mineralization inhibitory activity and are therefore expected tohave at least some of the same uses as the lower molecular weightpolypeptides. However, the most active materials for the purposes of thepresent invention, on a weight basis, are the lower molecular weightpolypeptides described herein.

The general structure of the polypeptides of the present invention is asfollows:

    poly (X).sub.n poly (Y).sub.m

where

each X independently is aspartic acid, glutamic acid, phosphoserine,phosphohomoserine, phosphotyrosine, or phosphothreonine,

each Y independently is alanine, leucine, isoleucine, valine, glycine orother nonpolar, amino acid residues,

n is 2 to 60, preferably 15-50, more preferably 30-50,

m is 2 to 60, preferably 3-15, more preferably 4-10,

n+m is ≧5, preferably n+m is 15-80, more preferably 15-40.

and salts thereof.

As can be seen from the general formula, the anionic amino acids areclustered on one end of the amino acid chain, whereas the nonpolar aminoacids are clustered on the other end. Thus, these polypeptides are notrandom copolymers as disclosed by several of the prior art referencesdiscussed above. In the formula, the X amino acids may either be at theC-terminus or the N-terminus. In other words, the aspartic acid,glutamic acid, etc. residues may be segregated at the N-terminal or theC-terminal.

The X amino acids may be entirely comprised of any one of the X group,or may be any combination of members of the group. Similarly, the Yamino acids may be entirely any one of the Y group, or may be anycombination of members of the group. For example, poly (X) could be madeup entirely of phosphorylated amino acids.

Salts of any of the acidic residues set forth above, especially thesodium and potassium salts, are also within the scope of this invention.When phosphorylated amino acids are incorporated in the compounds, theymay be present as salts, e.g., Ca⁺², Mg⁺², di-Na⁺, di-K⁺, and the like.Further, salts of the amino group, such as the p-toluenesulfonate,acetate, etc. are also contemplated.

Peptides wherein up to 10% of the X (anionic) residues are replaced by Y(non-polar) residues and vice versa are also within the scope of thisinvention. To illustrate this possibility, the following peptide isconsidered:

    H.sub.2 N--(Ala).sub.10 --(Asp).sub.10 --OH ,

The Y residues are ten alanines. One of these residues (10%) could bereplaced by an anionic residue (e.g., aspartic acid or glutamic acid).Similarly, the X residues are ten aspartic acid residues. One of thesecould be replaced by a non-polar amino acid (e.g., alanine, glycine,valine, etc.). Naturally, only integral numbers of replacement aminoacids are possible.

Specific preferred examples of formulas of polypeptides according to thepresent invention are the following:

    H.sub.2 N-(Asp).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --OH

    H.sub.2 N-(pSer).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(pSer).sub.n --OH

    H.sub.2 N-(Glu).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Glu).sub.n --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --(pSer).sub.x --OH

    H.sub.2 N-(Ala).sub.m --(Glu).sub.n --(pSer).sub.x --OH

wherein:

n=10-60, preferably 15-50.

m=2-10, preferably 3-8.

x=2-5, preferably 2-3.

[pSer=phosphoserine; that is, serine which has been phosphorylated onthe side chain hydroxyl].

In each of the above formulas, some or all of the alanine residues maybe replaced by other nonpolar amino acids, such as leucine, isoleucine,valine and glycine. Similarly, some or all of the aspartic acid residuesmay be replaced by other anionic amino acids such as glutamic acid, andvice versa. Further, some of the glutamic acid residues or aspartic acidresidues may be replaced by phosphoserine, phosphohomoserine,phosphotyrosine, phosphothreonine or other phosphorylated amino acids.Generally, amino acids containing a free hydroxyl group can bephosphorylated on the hydroxyl group. The phosphoserines could also bephosphohomoserine, phosphotyrosine, or phosphothreonine.

Some specific preferred embodiments of the present invention are thefollowing compounds:

    H.sub.2 N-(Ala).sub.5 --(Asp).sub.18 --(pSer).sub.2 --OH

    H.sub.2 N-(Ala).sub.5 --(Asp).sub.15 --OH

    H.sub.2 N-(Ala).sub.8 --(Asp).sub.40 --OH

As can be seen from the above description of the present compounds, alarge number of polypeptides fall within the scope of the presentinvention. However, each of them has in common the structural feature ofclustered hydrophobic or nonpolar amino acids on one end of thepolypeptide and clustered anionic amino acids on the other end of thepolypeptide. They are also generally small polypeptides having from10-80 amino acid residues, preferably 10-60 amino acid residues, mostpreferably 20-50 amino acid residues.

Methods of synthesis

The products of the invention may be synthesized by any number oftechniques now available for synthesis of simple and complex lowmolecular weight polypeptides. Generally speaking, these techniquesinvolve stepwise synthesis by successive additions of amino acids toproduce progressively larger molecules. The amino acids are linkedtogether by condensation between the carboxyl group of one amino acidand the amino group of another amino acid to form a peptide bond. Tocontrol these reactions, it is necessary to block the amino group of oneamino acid and the carboxyl group of the other. The blocking groupsshould be selected for easy removal without adversely affecting thepolypeptides, either by racemization or by hydrolysis of formed peptidebonds. Certain amino acids have additional functional groups such as thecarboxyl groups of aspartic acid and glutamic acid and the hydroxylgroups of serine, homoserine and tyrosine. It is usually necessary toblock these additional groups with an easily removed blocking agent, sothat they do not interfere with the desired condensation for theformation of peptide bonds.

A wide variety of procedures exist. for the synthesis of polypeptides,and a wide variety of blocking agents have also been devised. Most ofthese procedures are applicable to the peptides of the presentinvention. The preferred method for synthesis of the subject peptides isa solid-phase technique. In this procedure, an amino acid is bound to aresin particle, and the peptide is generated in a stepwise manner bysuccessive additions of protected amino acids to the growing chain. Thegeneral procedure is well known, and has been described in manyarticles, for example: Merrifield, R. B., J. Am. Chem. Soc. 96,2986-2993, 1964.

A preferred solid-phase method is described hereinbelow.

The peptides were made using automated, solid-phase peptide synthesis asdepicted in FIG. 1. The carboxy terminal amino acid was preloaded via anorganic linker (PAM, 4-oxymethyl phenylacetamidomethyl) covalentlyattached to an insoluble polystyrene resin cross-linked with divinylbenzene.

In this preferred embodiment, the C-terminus of the polypeptides wasaspartate and the C-terminal region polyanionic, with hydrophobicresidues added to the PH polypeptides on the N-terminus. This is theopposite of the structure of related natural protein inhibitors. Thereason for this orientation is that the aspartate-PAM linkage is easierto cleave at the end of the synthesis as compared to hydrophobic aminoacid-PAM linkages, resulting in greater yields. It is not likely thatthe positioning of the polyanionic or hydrophobic regions at the C vs.the N terminus matters with regard to activity.

The t-Boc strategy of protection of the terminal amine was used. Theside chain carboxyl group of aspartate and the OH of serine were bothprotected by O-benzyl groups. Final cleavage of the peptide from theresin and final R-group deprotection was achieved using hydrofluoricacid (HF) or trifluoromethyl sulfonic acid (TFMSA) according toestablished procedures (Bergot, B. J. and S. N. McCurdy, AppliedBiosystems Bulletin (1987)). An automated peptide synthesizer (AppliedBiosystems, model 430-A) was used. A routine synthesis produced 0.5mmole of peptide-resin. Yield after cleavage and purification wasapproximately 60 to 70%.

Purification was accomplished by recrystallizing the peptides from anorganic solvent such as methyl-butyl ether, then dissolving the crystalsin a small amount (e.g. 5.0 ml) of distilled water in a dialysis bag (MWcutoff 500 daltons), dialysis for removal of residual solvents, andlyophilization. Purity of the peptides was checked by reversed-phaseliquid chromatography (Varian 5560) using a C-18 column and 0.1 %trifluoroacetic acid and acetonitrile as solvents, or by gel permeationusing a column for separations between 1000 and 30,000 daltons with 0.05M trishydroxymethyl amino methane (pH 8.0) as the mobile phase. Thisapproach yielded peptides of precisely-known sequence and molecularsize.

Alternatively, polyanionic/hydrophobic peptides of approximately-knownsequence and size may be made by conventional thermal polymerization ofthe polyanionic region and the hydrophobic regions separately usingR-group protected amino acids (Melius, P. and J.Y.P. Sheng, Bioorg. Chem4, 385 (1975)). Next the polyanionic and hydrophobic regions can belinked thermally, followed by deprotection of the R-groups usingcleavage reagents. There is some evidence to suggest that apolyanionic/hydrophobic peptide may assemble naturally under thermalpolymerization conditions, without the need for separate synthesisfollowed by attachment of the two regions (Phillips, R. D. and P.Melius, Int. J. Peptide Protein Res. 6, 309-319 (1974)).

A preferred embodiment of the peptide inhibitors involves theincorporation of 2 or more phosphoserine residues at any location in thepolyanionic region of the molecule. This idea is consistent with theknown activity of simple phosphate and polyphosphates as inhibitors ofcrystallization and the similar activity of phosphonates. In addition,the inventors' work and recent reports (Hay, D. I. et al, Calcif. TissueInt. 40, 126-132 (1987)) suggest that the presence of phosphoserineresidues in natural proteins significantly enhances inhibitory activity.Therefore, peptides containing phosphoserine have been preparedaccording to the method of Riley et al., J. Am. Chem. Soc. 1957,1373-1379.

This method involves derivitization of serine residues after peptidesynthesis using diphenyl phosphochloridate in the ratio of X molesserine to X+1 moles of the phosphochloridate at room temperature for 2hours with dimethylformamide as solvent. This produces a diphenylphosphate ester plus polypeptide that can be recrystallized from ether.The phosphopeptide ester is then dissolved in 0.1 M NH₄ HCO₃ and thephenyl groups removed by reduction by sparging with H₂ gas in thepresence of a platinum or palladium oxide catalyst. The phosphopeptideis then recrystallized and purified. Other useful derivatives, forexample, sulfated, phosphonated, and sulfonated peptides are alsocontemplated. In each case, a phosphate moiety could be replaced with asulfate, phosphonate or sulfonate group.

Activity Assays

To measure the ability of the peptides of the present invention toinhibit mineralization, a number of assays have been developed. Theseassays include the following:

1. pH--drift assay - CaCO₃

2. pH--drift assay calcium phosphate.

3. pH--stat assay - CaCO₃

4. constant composition assay - CaCO₃.

The following examples describe how these various assays have beenemployed to measure the ability of the polypeptides to inhibitmineralization.

EXAMPLE 1: The pH-Drift Assay - Calcium Carbonate

A solution supersaturated with respect to CaCO₀₃ is prepared byseparately pipetting 0.3 ml of 1.0 M CaCl₂ dihydrate and 0.6 ml of 0.5 MNaHCO₃ into 29.1 ml of artificial seawater (0.5 NaCl, 0.011 M KCl). Thereaction vessel is a 50 ml, 3-necked, round-bottom flask partiallyimmersed in a thermostated water bath at 20° C. The reaction vessel isclosed to the atmosphere to minimize exchange of CO₂ The reaction isstarted by adjusting the pH upward to 8.3 by titration of μl amounts of1 N NaOH by digital pipette. The initial concentrations are 10 mM eachof Ca²⁺ and dissolved inorganic carbon (DIC). The reaction is monitoredby pH electrode and recorded by strip chart.

After a period of stable pH during which crystal nuclei form, the pHbegins to drift downward until the reaction ceases due to depletion ofreactants and the lowering of pH. The reaction may be quantified bycalculations based on DIC equilibria to give the amount of precipitationversus time. In FIG. 2 it can be seen that a change in pH is directlyproportional to a change in DIC from pH 8.3 to 7.7, but below 7.7 thebuffering effect of the DIC system leads to greater changes in DIC perunit pH.

EXAMPLE 2: The pH-drift assay: Calcium Phosphate

A solution supersaturated with respect to calcium phosphate is preparedby separately pipetting 0.1 ml of 1.32 M CaCl₂ dihydrate and 0.1 ml of0.90 M NaHl.sub. PO₄ into 29.8 ml of distilled water. This yieldsinitial concentrations of 4.4 mM Ca²⁺ and 3.0 mM dissolved inorganicphosphorus (DIP). The reaction vessel is a 50 ml, round-bottom, 3-neckedflask partially immersed in a thermostated water bath at 20° C. Thereaction vessel is closed to the atmosphere. The reaction begins uponmixing the reactants with an initial pH of 7.4.

FIG. 3 shows data obtained from this type of assay. Amorphous calciumphosphate (ACP) nucleates immediately and slowly grows as indicated by aslight decrease in pH during the first 30 minutes or so of the assay.Following this, ACP begins to transform to calcium hydroxylapatite(HAP), Ca₁₀ (PO₄)₆ (OH)₂, as indicated by a marked acceleration in thedownward pH drift. The reaction ceases as reactants are depleted and thepH is lowered.

EXAMPLE 3: The pH-stat assay

The following description relates to FIG. 4.

A. The effect on CaCO₃ crystallization of various concentrations ofEDTA-extracted soluble matrix from oyster shell were compared in thepH-stat assay. The medium consisted of 25 ml 0.5 M NaCl, 0.01 M KCl, and10 mM DIC with the initial pH adjusted to 8.5. The reaction vessel wasthermostated at 25° C. To initiate crystal growth, CaCl₂ was added to afinal concentration of 10 mM (first arrows). After an initial decreasein pH to 8.3, probably due to ion-pairing, and an induction period ofapproximately 2 minutes, crystals appeared in solution.

The crystal growth is proportional to the amount of 0.5 M NaOH titrantrequired to maintain a constant pH during the overall reaction Ca²⁺+HCO₃ ⁻ →CaCO₃ ^(+H) ⁺. The pH was held constant by autotitration.

B. After crystals grew to an equivalent of 25 μmol of titrant, variousquantities of matrix were added (second arrow) ranging from 0.5 μg/ml(a) to 1.5 μg/ml (d) final concentration. Note that the rate of crystalgrowth after addition of matrix decreased relative to the control rateas matrix concentration increased, and growth was terminated at 1.5μg/ml.

C. For comparing various inhibitors (for example, see Table 1), thegrowth rate immediately following the addition of inhibitor iscalculated as a percent of the growth rate of the crystals before theaddition of inhibitor. From examining the effects of a series ofconcentrations, the quantity of inhibitor required to reduce the controlrate of crystal growth by 50% can be determined graphically.

EXAMPLE 4: The Constant Composition Assay: The Effect of Oyster ShellMatrix, Polyaspartate, and HEDP on the rate of CaCO₃ crystal growth

A 50 ml solution of 10 mM CaCl₂ and 10 mM dissolved inorganic carbon(DIC) was adjusted to pH 8.34 by digital pipetting of μl amounts of 1 NNaOH. The temperature was held constant at 20° C. using a water-jacketedreaction vessel and a thermostated bath. After about a 6 minute periodduring which CaCO₃ nucleation occurs, the pH beings to drift downward asCa²⁺ and CO₃ ²⁻ ions are removed from solution as solid crystals. At pH8.30, inhibitors were added which stabilized the pH drift for certainperiods of time. The concentrations of inhibitors used in theexperiments shown in FIG. 5 were matched so that the amount ofstabilization or inhibition of nucleation was the same, in this caseabout 2 hours. The doses required for this effect ranged between 0.1 to0.5 μg/ml of inhibitor depending on the type When crystallization beganagain, the pH would drift downward. However, it was held constant at pH8.30±0.02 by automatic titration of 0.5 M CaCl₂ and 0.5 M NaCO₃ fromseparate burettes linked via the pH electrode to a computer-assistedtitrimeter.

In FIG. 5, notice the dramatic acceleration of the rate ofcrystallization in the presence of hydroxyethylidene diphosphonate(HEDP), perhaps the most commonly-used CaCO₃ inhibitor for watertreatment. This effect was not observed in the presence of matrix or thematrix analog over the time periods studied. These results suggest quitedifferent mechanisms of action for the two classes of crystallizationinhibitors.

EXAMPLE 5: The effects of synthetic peptides on CaCO₃ crystallization

The calcium carbonate pH-drift assay was used to assay three peptides,peptides 1, 2 and 3. Peptide 1 =poly (Asp)₂₀, peptide 2=poly (Asp-Gly)₇(Ala)₆, and peptide 3=poly (Asp)₁₅ (Ala)₅. The conditions of the assaywere 10 ml Ca, 10 ml dissolved inorganic carbon at 20° C. and 30 ml ofartificial seawater (0.5 M NaCl, 0.01 M KCl). The results are shown inFIG. 6.

Peptide 3, a PH peptide, exhibited significantly enhanced inhibition ofcrystal nucleation, the formation of new crystals in solution. Theeffect on nucleation is seen in the increased duration of the inductionperiod of stable pH prior to the period of crystal growth when pH driftsdownward. No effect on crystal growth (the increase in size ofpre-existing crystals) was observed in this assay, as shown by the slopeof the recorder tracing during the phase of crystal growth. The activityof the PH peptide was compared to that of a polyaspartate of equivalentchain length (peptide 1) at equal doses. It was also interesting to notethat an ordered copolymer of aspartate-glycine with a hydrophobic tailof alanine residues (peptide 2) has very little activity relative to thecontrol curve. The repeating Asp-Gly arrangement is the structurepredicted for acidic proteins from mollusk shells based on publishedreports (e.g., Weiner, S., Biochemistry 22, 4139-4145 (1983)).

EXAMPLE 6: Inhibition of CaCO₃ crystallization by synthetic peptides:pH-drift assay

A more detailed comparison of effects as seen in Example 5 is presentedin Table 1. The PH peptide H--(Ala)₅ --(Asp)₁₅ --OH exhibited thegreatest inhibition of crystal nucleation. Again, there was no cleareffect on crystal growth using this assay. The relative lengths of thepolyanionic and hydrophobic regions are important to inhibitory activityas shown by decreased activity of the PH peptide H--(Ala)₁₀ --(Asp)₁₀--OH.

The length of the polyanionic region that provides the greatest affinityfor crystal surfaces falls in the range of (Asp or Glu) ₁₅ to (Asp orGlu)₆₀, with preferred lengths in the range (Asp or Glu)₃₀ to (Asp orGlu)₅₀. The preferred length of the hydrophobic region falls in therange of 3 to 8, e.g. (Ala)₃ to (Ala)₈.

Peptides that might have been predicted to have high activity (the last4 peptides) based on published reports about the structure of naturalinhibitory proteins did not exhibit enhanced activity relative to theperformance of polyaspartate or the more active PH peptide. These last 4peptides are ordered copolymers of aspartate, glycine, and serine, inone case having a polyalanine tail.

                                      TABLE 1                                     __________________________________________________________________________    INHIBITION OF CaCO CRYSTALLIZATION BY SYNTHETIC PEPTIDES                      pH-DRIFT ASSAY                                                                                 CONCENTRATION                                                                            INDUCTION                                                                             CRYSTAL GROWTH                            PEPTIDE          μg/ml   PERIOD, MIN                                                                           RATE, pH/MIN                              __________________________________________________________________________    CONTROL          --         7.1 ± 1.96                                                                         0.048 ± 0.0041                         H--(ASP)20--OH   0.02       21.5 ± 0.50                                                                        0.047 ± 0.0030                         (polyaspartate)  0.035      39.2 ± 10.8                                                                        0.041 ± 0.0050                                          0.050      >180    --                                        H--(ALA).sub.5 --(ASP).sub.15 --OH                                                             0.005      11.3 ± 3.37                                                                        0.047 ± 0.0025                                          0.01       35.4 ± 13.8                                                                        0.040 ± 0.0034                         H--(ALA).sub.10 (ASP).sub.15 --OH                                                              0.050      32.0 ± 10.4                                                                        0.042 ± 0.0039                         H--(GLY--ASP).sub.10 --OH                                                                      0.10       36.0 ± 5.20                                                                        0.047 ± 0.0097                         H--(ALA).sub.6 --(GLY-13 ASP).sub.7 --OH                                                       0.20       7.2 ± 0.83                                                                         0.048 ± 0.0042                         H--(SER--ASP).sub.10 --OH                                                                      0.1        29.8 ± 4.99                                                                        0.048 ± 0.0040                         H--(GLY--SER--ASP).sub.7 --OH                                                                  0.1        5.83 ± 1.04                                                                        0.057 ± 0.0034                         __________________________________________________________________________

EXAMPLE 7: Inhibition of CaCO₃ crystal growth by synthetic peptides:constant composition/seeded crystal assays

In this assay, 0.1 ml of 10 M CaCl₂ and 0.2 ml of 0.5 M NaHCO₃ are addedto 49.7 ml of artificial seawater. To start the reaction, 0.1 ml of anaqueous slurry of CaCO₃ crystals (Baker Analytical Reagents) is added,and the pH is adjusted to 8.5 by titration of μliter amounts of lN NaOH.Crystals begin to grow immediately in the solution, which is 2.0 mM Ca²⁺and 1.6 mM dissolved inorganic carbon. The reaction again is monitoredby a pH electrode linked to a computer assisted titrator set to maintainthe pH at 8.50. The titrator simultaneously adds 0.1 M CaCl₂ and 0.1 MNa₂ CO₃ (pH 11.0) to replace the Ca²⁺ and CO₃ ²⁻ that are removed fromsolution as CaC03 crystals grow. Thus, the chemical potential of thesolution is kept constant. When inhibitors are used, they are addedafter the addition of the calcium. The experiments shown in FIG. 7represent average results for at least triplicate assays for eachmolecule. Control curves are run before or after each set ofexperimental curves. This assay reveals clearer effects of inhibitors oncrystal growth because growth can be monitored under constant conditionsover extended periods. Effects of inhibitors on crystal nucleation, onthe other hand, are not seen in this assay because there is no inductionperiod prior to crystal growth during which nucleation occurs.Conversely, the pH-drift assay is best for measuring effects onnucleation as shown by changes in the induction period. But the pH-driftassay is not good for measuring crystal growth because the reaction isself-limiting during crystallization due to depletion of reactants andlowering of pH.

In these experiments, all of the polypeptides were added to the mediumof crystal growth at a dose of 0.02 μg/ml prior to the addition of 2.5mg of CaCO₃ seeds. The specific effect of increased inhibition ofcrystal growth was seen as the length of the polyalanine tail wasincreased to 10 residues with the polyaspartate region kept at 15residues. For comparative purposes, polyaspartate molecules ofincreasing chain lengths were also tested.

The results suggest that the most active molecules on a weight basiswill have a specific chain length of the polyanionic region to provideoptimum affinity for crystal surfaces. In addition, a specific chainlength of the hydrophobic region is required, presumably to provideoptimum coverage of the crystal surface including the zone immediatelysurrounding the surface.

EXAMPLE 8: The effect of some inhibitors on calcium phosphate formation

The data in Table 2 show that CaCO₀₃ crystallization inhibitory activityof a molecule can also be applied to calcium phosphate formation. Boththe oyster shell protein and polyaspartate can inhibit both CaCO₃ andcalcium phosphate formation. The presence of phosphorylated residues inthe oyster shell protein enhances the inhibitory activity with respectto amorphous calcium phosphate (ACP) formation and apatite formation.

The effects of some other molecules are shown for comparative purposes.The phosphino-carboxylate copolymer and polymaleate are representativeof industrial polymers currently in use for prevention of mineralscaling in cooling towers and other equipment. The phosphonatediethylene triamine pentamethylene phosphonic acid (DENTPP), is the mosteffective phosphonate known for inhibition of calcium phosphateformation. The salivary protein statherin is the most effectiveinhibitor among a group of salivary proteins that regulate mineraldeposition in the oral cavity.

                                      TABLE 2                                     __________________________________________________________________________    EFFECT OF SOME INHIBITORS ON CALCIUM PHOSPHATE FORMATION.sup.1                          PERIOD OF  pH/min,    pH/min,                                       INHIBITOR ACP FORMATION                                                                            ACP FORMATION                                                                            APATITE FORMATION                             __________________________________________________________________________    None (control,                                                                          31.3 ± 4.36 min                                                                       0.0047 ± 00.23                                                                        0.090 ± 0.013                              n = 20 ± S.D.)                                                             Oyster shell                                                                            65 minutes 0.0030     0.068                                         matrix, phosphor-                                                             ylated, 10 μg/ml.sup.2                                                     Oyster shell                                                                            40 minutes 0.0034     0.105                                         matrix dephosphory-                                                           lated, 10 μg/ml                                                            Polyaspartate,                                                                          30 minutes 0.0040     0.106                                         MW 15,000,                                                                    10 μg/ml                                                                   Polyaspartate,                                                                          48 minutes 0.0050     0.070                                         MW 15,000,                                                                    15 μg/ml                                                                   Phosphino-carboxy-                                                                      69 minutes 0.0025     0.052                                         late copolymer,                                                               10 μg/ml                                                                   Polymaleate,                                                                            83 minutes 0.0014     0.051                                         10 μg/ml                                                                   DENTPP,   100 minutes                                                                              0.0012     0.019                                         phosphonate,                                                                  5 μg/ml                                                                    Statherin.sup.3                                                                         48 minutes 0.0045     0.0090                                        37 μg/ml                                                                   __________________________________________________________________________     .sup.1 Assay Conditions: Ca at 4.4 mM, dissolved inorganic phosphorus at      3.0 mM, initial pH 7.40, 20° C.                                        .sup.2 Results for experimental molecules are given as the average values     for at least triplicate experiments.                                          .sup.3 Adapted from Hay, D. I., E. L. Moreno, and D. H. Schlesinger. 1979     Phosphoproteininhibitors of calcium phosphate precipitation from salivary     secretions. Inorganic Perspectives in Biology and Medicine 2, 271-285.   

EXAMPLE 9: The effect of phosphorylation on inhibition of CaCO₃formation by oyster shell matrix

The purpose of the measurements reported in Table 3 was to clarify theinfluence of phosphorylation of the oyster shell protein on its activitywith respect to CaCO₃ formation. Again, it is clear that the presence ofphosphorylated residues leads to significantly enhanced inhibitoryactivity.

                  TABLE 3                                                         ______________________________________                                        EFFECT OF PHOSPHORYLATION ON INHIBITION OF                                    CaC0.sub.3 FORMATION.sup.l BY OYSTER SHELL MATRIX.sup.2                                      Pi     I.sub.50                                                               % by wt.                                                                             μg ml.sup.-1                                         ______________________________________                                        Untreated        13.6-15.8                                                                              1.4-2.2                                                              (N = 8)  N = 8)                                              Partial dephos-  7.2- 10.0                                                                              2.5- 3.4                                            phorylated.sup.3 (N = 3)  (N = 3)                                             Dephosphorylated 0.6-3.7  6.4-8.3                                                              (N = 4)  (N = 3)                                             ______________________________________                                         .sup.1 Assay conditions: 10 mM Ca, 10 mM dissolved inorganic carbon, 25 m     artificial seawater, pH 8.3, pHstat assay.                                    .sup.2 The protein from oyster shell was solubilized by dissolution of        crushed shell in 10% EDTA, pH 8.0, followed by isolation using gel            permeation (sephacryl) to yield a fraction of about 30,000 daltons that       was then concentrated by tangential flow dialysis, lyophilized, and           frozen.                                                                       .sup.3 Dephosphorylation was accomplished by treatment with alkaline          phosphatase according to Termine, J. D. and K. M. Conn, 1976, Inhibition      of apatite formation by phosphorylated metabolites and macromolecules,        Calcif. Tiss. Res. 22, 149-157. Phosphorus was measured                       spectrophotometrically according to Marsh, B. B., 1959, The estimation of     inorganic phosphate in the presence of adenosine triphosphate. Biochem.       Biophys. Acta 32, 351-361.                                               

Use of the Present Polypeptides

The present materials serve as inhibitors of dental tartar (calculus)and plaque formation (referred to herein as tartar barrier agents) forhuman or animal use. The oral compositions according to this inventionmay comprise any conventional pharmaceutically acceptable oral hygieneformulation that contains and is compatible with an effective amount ofan antidental calculus agent as disclosed herein. Such formulationsinclude, for example, mouthwashes, rinses, irrigating solutions,abrasive and non-abrasive gel dentifrices, denture cleansers, coateddental floss and interdental stimulator coatings, chewing gums,lozenges, breath fresheners, foams and sprays. These formulations may beused to treat natural or artificial tooth enamel, or any orallycompatible material which is subject to mineral deposition. Althoughhuman use is preferred, use in animals is also possible.

The tartar barrier agents may be present in the formulations ineffective concentrations generally in the range of from about 0.05 wt. %to as much as 30 wt. % or the limit of compatibility with a vehicle. Apreferred concentration range for the agents of the formulations of theinvention is from about 0.5 to about 10 wt. %. A more preferred range isfrom about 2 to about 8 wt. %.

The pH of these preparations should be between about pH 5 and 10,preferably between pH 5 and 8, more preferably between about 6 and 7.5.A pH lower than 5 is undesirable because of the possible enhancement ofenamel demineralization.

Suitable conventional pharmaceutically acceptable vehicles that can beemployed with the tartar barrier agents to prepare the compositions ofthis invention may comprise water, ethanol; such humectants aspolypropylene glycol, glycerol and sorbitol; such gelling agents ascellulose derivatives, for example, Methocel carboxymethylcellulose (CMC7MF) and Klucel HF, polyoxypropylene/polyoxyethylene block copolymers,for example, Pluronic F-127, Pluronic F-108, Pluronic P-103, PluronicP-104, Pluronic P-105, and Pluronic P-123, colloidial magnesiumaluminosilicate complexes such as Veegum, and mucoprotein thickeningagents such as Carbopol 934; gel stabilizers such as the silicondioxides, for example Cab-0-Sil M5, and polyvinylpyrrolidone; sweetenerssuch as sodium saccharin and aspartame; preservatives such as citricacid, sodium benzoate, cetylpyridinium chloride, potassium sorbate,methyl and ethyl parabens; detergents such as sodium lauryl sulfate,sodium cocomonoglyceride sulfonate, sodium lauryl sarcosinate andpolyoxyethylene isohexadecyl ether (Arlasolve 200) and approved colorsand flavors.

The following specific examples will serve further to illustrate thetartar barrier agent compositions of this invention.

    ______________________________________                                        EXAMPLE A - Mouthwash Solution                                                Tartar barrier agent    0.5-2.0% w/w                                          Glycerol (Humectant)    6.0                                                   Pluronic F-108          1.0                                                   Sodium saccharin (sweetener)                                                                          0.3                                                   Deionized Water         q.s.                                                  Flavors                 1.0                                                                           100.0                                                 EXAMPLE B - Mouthwash Solution                                                Tartar barrier agent    0.5-3.0% w/w                                          Ethanol, USP            15 0                                                  Pluronic F-108 (foaming agent)                                                                        2.0                                                   Glycerol (humectant)    10.0                                                  Sorbitol (humectant)    10.0                                                  Sodium saccharin (sweetener)                                                                          0.2                                                   Deionized Water         q.s.                                                  Flavors                 0.2                                                                           100.0                                                 EXAMPLE C - Abrasive Dentrifice Gel                                           Tartar barrier agent    2.0-10.0% w/w                                         Fumed Silica (abrasive) 55.0                                                  Sodium Lauryl Sulfate (detergent)                                                                     1.5                                                   Glycerol (Humectant)    10.0                                                  Carboxymethylcellulose  2.0                                                   (gelling agent)                                                               Sodium saccharin (sweetener)                                                                          0.2                                                   Sorbitol (humectant)    10.0                                                  Flavors                 1.0                                                   Deionized Water         q s.                                                  Preservative            0.05                                                                          100.00                                                EXAMPLE D - Chewing Gum                                                       Tartar barrier agent    1 0-11.0% w/w                                         Gum Base                21.3                                                  Sugar                   48.5-58.5                                             Corn Syrup (Baume 45)   18.2                                                  Flavors                 1.0                                                                           100.00                                                EXAMPLE E - Nonabrasive Gel Dentrifice                                        Tartar barrier agent    0.05-30.0% w/w                                        Sorbistat (preservative)                                                                              0.15                                                  Deionized Water         q.s.                                                  Silicon Dioxide (gel stabilizer)                                                                      1.0                                                   Pluronic F-127 (gelling agent)                                                                        20.0                                                  Sodium saccharin        0.2                                                   Flavors                 1.5                                                                           100.0                                                 ______________________________________                                    

In addition to the above material which can be included in the presenttartar barrier compositions, it is also contemplated to include thereina protease inhibitor to prevent the present peptides and polypeptidesfrom being degraded by various proteolytic enzymes.

Examples of such inhibitors include aprotinin and trypsin inhibitortypes I-P, I-S, II-L, II-O, II-S, II-T, III-O, and IV-O, although otherinhibitors are within the scope of this invention. Similarly, whenphosphopeptides are employed, it is contemplated to use phosphataseinhibitors in conjunction with the polypeptide to prevent or inhibitdephosphorylation of the polypeptides. Examples of such phosphataseinhibitors are sodium fluoride, adenosine diphosphate, and vinylether/maleic acid polymers (gantrez). Use of other phosphataseinhibitors is also possible.

The present peptides and polypeptides could also be linked toantibodies, particularly those against cavity-causing bacteria, or theantibodies could be added to a tartar barrier composition to enhanceantibacterial activity.

The invention now being fully described, it will be apparent to one ofordinary skill in the art that many changes and modifications can bemade thereto, without departing from the spirit or scope of theinvention as set forth therein.

What is claimed as new and desired to be secured by Letters Patent ofthe United States is:
 1. A method of treating a tooth so as to inhibitdeposition of mineral thereon, which comprises contacting said toothwith a mineral deposition inhibitory effective amount of a poly-aminoacid compound which has the clustered polyanionic/clustered non-ionic,partly hydrophobic structure:

    poly (X).sub.n poly (Y).sub.m

where each X is a residue independently selected from the groupconsisting of aspartic acid, glutamic acid, phosphoserine,phosphohomoserine, phosphotyrosine, and phosphothreonine, each Y isindependently a residue selected from the group consisting of alanine,leucine, isoleucine, valine and glycine,

    n is 2 to 60,

    m is 2 to 60, and

    n +m >5,

and wherein poly (X)_(n) may contain up to 10% of the Y residues andpoly (Y)_(m) may contain up to 10% of the X residues, and salts thereof,in combination with an orally acceptable vehicle compatible with saidcompound.
 2. A method according to claim 1, in which the first X residuein the poly-amino acid compound is at the N-terminal of said compoundand the last Y residue is at the C-terminal of said compound.
 3. Amethod according to claim 1, in which the first X residue in thepoly-amino acid compound is at the C-terminal of said compound and thelast Y residue is at the C-terminal of said compound.
 4. A methodaccording to claim 1, wherein the poly-amino acid compound has a formulaselected from the group consisting of:

    H.sub.2 N-(Asp).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --OH

    H.sub.2 N-(pSer).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(pSer).sub.n --OH

    H.sub.2 N-(Glu).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Glu).sub.n --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --(pSer).sub.x --OH, and

    H.sub.2 N-(Ala).sub.m --(Glu).sub.n --(pSer).sub.x --OH

wherein: n=10-60, m=2-10, and x=2-5.
 5. A method according to claim 4,wherein the poly-amino acid compound is: H₂ N-(Ala)₅ --(Asp)₁₈ --(pSer)₂--OH.
 6. A method according to claim 4, wherein the poly-amino acidcompound is: H₂ N-(Ala)₅ --(Asp)₁₅ --OH.
 7. A method according to claim4, wherein the poly-amino acid compound is: H₂ N-(Ala)₈ --(Asp)₄₀ --OH.8. A method according to claim 1, wherein in the formula for thepoly-amino acid compound, n=20-40, m=2-8, and the number ofphosphorylated amino acids is 0-3.
 9. A method according to claim 1,wherein said aspartic acid and glutamic acid residues in said poly-aminoacid compound are in the form of sodium or potassium salts.
 10. A methodaccording to claim 1, wherein said phosphoserine, phosphohomoserine,phosphotyrosine, and phosphothreonine residues in said poly-amino acidcompound are in the form of disodium, dipotassium, calcium or magnesiumsalts.
 11. A method according to claim 1, wherein said tooth is that ofa human.
 12. A dentifrice composition, which comprises a mineraldeposition inhibitory effective amount of a poly-amino acid compoundwhich has the clustered polyanionic/clustered non-ionic, partlyhydrophobic structure:

    poly (X).sub.n poly (Y).sub.m

where each X is a residue independently selected from the groupconsisting of aspartic acid, glutamic acid, phosphoserine,phosphohomoserine, phosphotyrosine, and phosphothreonine, each Y isindependently a residue selected from the group consisting of alanine,leucine, isoleucine, valine and glycine, n is 2 to 60, m is 2 to 60, andn+m≧5, and wherein poly (X)n may contain up to 10% of the Y residues andpoly (Y)m may contain up to 10% of the X residues, and salts thereof, incombination with an orally acceptable dentifrice composition compatiblewith said compound.
 13. A composition according to claim 12, in the formof an oral hygiene formulation selected from the group consisting ofmouthwashes, rinses, irrigating solutions, abrasive gel dentifrices,nonabrasive gel dentrifices, denture, cleansers, coated dental floss,interdental stimulator coatings, chewing gums, lozenges, breathfresheners, foams and sprays.
 14. A dentifrice composition according toclaim 12, in which the first X residue in the poly-amino acid compoundis at the N-terminal of said compound and the last Y residue is at theC-terminal of said compound.
 15. A dentrifrice composition according toclaim 12, in which the first X residue in the poly-amino acid compoundis at the C-terminal of said compound and the last Y residue is at theN-terminal of said compound.
 16. A dentrifrice composition according toclaim 12, wherein the poly-amino acid compound has a formula selectedform the group consisting of:

    H.sub.2 N-(Asp).sub.n --(ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --OH

    H.sub.2 N-(pSer).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m -(pSer)n-OH

    H.sub.2 N-(Glu).sub.n --(Ala).sub.m --OH

    H.sub.2 N-(Ala).sub.m --(Glu).sub.n --OH

    H.sub.2 N-(Ala).sub.m --(Asp).sub.n --(pSer).sub.x --OH, and

    N.sub.2 N-(Ala).sub.m --(Glu).sub.n --(PSer).sub.x --OH

wherein: n=10-60, m=2-10, and x=2-5.
 17. A dentifrice compositionaccording to claim 16, wherein the poly-amino acid compound is:

    H.sub.2 N-(Ala).sub.5 --(Asp).sub.18 --(pSer).sub.2 --OH.


18. A dentifrice composition according to claim 16, wherein thepoly-amino acid compound is: H₂ N-(Ala)₅ --(Asp)₁₅ --OH.
 19. Adentifrice composition according to claim 16, wherein the poly-aminoacid compound is: H₂ N-Ala)₈ --(Asp)₄₀ --OH.
 20. A dentifricecomposition according to claim 16, wherein in the formula for thepoly-amino compound, n=20-40, m=2-8, and the number of phosphorylatedamino acids is 0-3.
 21. A dentifrice composition according to claim 12,wherein said aspartic acid and glutamic acid residues in said poly-aminoacid compound are in the form of sodium or potassium salts.
 22. Adentrifrice composition according to claim 12, wherein saidphosphoserine, phosphohomoserine, phosphotyrosin, and phosphothreonineresidues in said poly-amino acid compound are in the form of disodium,dipotassium, calcium or magnesium salts.